Traumatic brain injury (TBI) or traumatic neuronal injury (TNT) in vitro causes neuronal apoptosis, in part, through activation of caspase-3-like proteases. inhibition of caspase-3 in vitro reduces posttraumatic cell death and provides additive neuroprotection to that produced by agents that inhibit necrotic cell death. Caspase-3 activation is modulated by upstream caspases, including caspase-9 (intrinsic pathway) and caspase-8 (extrinsic pathway). Our preliminary data suggests that the caspase-9 pathway appears to be more important in neurotrauma. Akt (protein kinase B) is a well-established anti-apoptotic factor, which may act, in part, by modulating caspase activation. Akt itself can also be modulated by several factors, including the novel tumor suppression protein PTEN. Recent experiments in our laboratory have suggested a role for PTEN in neuronal apoptosis. We propose to examine the role of Akt in neuronal apoptosis after TB! and TNT, and elucidate the critical upstream and downstream signal transduction pathways involved. Specific hypotheses include: 1) caspase-9, but not caspase-8, represents an important upstream modulatory mechanism for caspase-3 mediated apoptosis after trauma; 2) Akt plays an important modulatory role in apoptosis following TBI or TN! in vitro; 3) Anti-apoptotic actions of Akt include inhibition of caspase-3 activation by phosphorylating the pro-apoptotic factor BAD, as well as through other non-caspase mechanisms; 4) The recently identified tumor suppressor factor PTEN is activated after trauma or trophic withdrawal and contributes to neuronal apoptosis, in part, by downregulating Akt activity; and 5) Development of PTEN antagonists may provide a novel neuroprotective treatment strategy for CNS injury. We propose the following specific aims: 1) To examine the relative contributions of the intrinsic (caspase-9) and extrinsic (caspase-8) pathways in modulating caspase-3-induced apoptosis in TB! and TNT, using complementary in vivo and in vitro model systems; 2) To establish an anti-apoptotic role for Akt in TBI and TNT and examine proposed mechanisms, including phosphorylation of BAD. 3) To demonstrate a pro-apoptotic role for PTEN in TBI and TNI, and show that this action results substantially from its ability to downregulate Akt, resulting in activation of caspase-9 and caspase-3; and 4) To show that PTEN antagonists, newly developed by us in collaboration, are neuroprotective following injury in vivo and in vitro.